Dilated and ?burnt out? hypertrophic cardiomyopathies are common genetic cardiomyopathies that lead to heart failure. Currently over 115,000 Veterans annually receive care for heart failure from the VA Health Care System. Despite efforts to implement guideline-directed medical therapy, the overall 5 year mortality is ~50% after diagnosis, so clearly this is a disease important to not only Veterans but also the general population. Myosin heavy chain 7 (MYH7) mutations are common causes of hypertrophic and dilated cardiomyopathies. Genetic testing for MYH7 variants have been limited by frequent identification of variants of unknown significance and the lack of disease-modifying therapies when pathogenic variants are identified. This proposal will identify MYH7 variants that will cause contractile dysfunction, the first step to the development of heart failure, and study the disease pathogenesis in human induced pluripotent stem cell-derived cardiomyocytes. Mutations in either the MYH7 S2 domain or the C1C2 domain of cardiac myosin binding protein C (cMyBPC) that disrupt the normal protein-protein interaction between S2/C1C2 have recently been shown to induce heart failure with reduced ejection fraction. This leads to the hypothesis that a subset of MYH7 mutation-induced cardiomyopathies are due to impaired interaction between these two proteins. The proposed work uses saturation mutagenesis and high-throughput modified yeast two-hybrid assays to identify nearly all mutations in the MYH7 S2 domain that disrupt normal protein-protein interaction with the C1C2 domain of cMyBPC. This will assist in identifying all clinically relevant MYH7 S2 variants that are susceptible to developing heart failure and generate a ?look up? table that would enable the confident identification of patients that could benefit from therapeutic intervention (Aim 1). Abnormally functioning mutant MYH7 protein raises the possibility of increased myosin degradation. This is supported by recent work demonstrating an upregulation of muscle RING-finger protein-1 (MuRF1), an E3 ligase that targets MYH7 and other sarcomeric proteins for degradation, in human induced pluripotent stem cell-derived cardiomyocytes expressing the pathogenic MYH7 E848G variant. This leads to the hypothesis that MuRF1 upregulation in MYH7 mutation-induced cardiomyopathies contributes to systolic dysfunction and that reducing MuRF1 levels will increase contractility. The proposed work will use gain-of-function and loss-of-function experiments to elucidate the role of MuRF1 in MYH7 mutation-induced cardiomyopathies (Aim 2). If successful it will determine if MuRF1 can be a novel therapeutic target for these genetic cardiomyopathies. The proposed work uses several innovative techniques. It combines cutting-edge high-throughput functional assays with mechanistic studies in genetically-edited human induced pluripotent stem cell-derived cardiomyocytes to identify patients with MYH7 mutations that are at risk of developing heart failure and then determines the suitability of a potential novel disease-modifying intervention. The high-throughput assays will integrate well with the proposed training in computational genomics. The method in Aim 1 can later be applied to interactions between MYH7 and other sarcomeric proteins to potentially identify all clinically relevant MYH7 variants. The mechanistic studies in human induced pluripotent stem cells in Aim 2 will help elucidate the pathogenesis of MYH7 mutation-induced cardiomyopathies and together with Aim 1 will form the basis for a future Merit Award proposal during the 4th year of the CDA2 award. Overall, the CDA2 will provide the additional training necessary for the nominee to use iPSC-based disease modeling with computational genomics to discover new disease-modifying therapies with precision medicine approaches.